216: 53BP1-RIF1 and DNA-PKcs: distinct interactions in chromosomal break repair

Makins K et al., Nature Communications - This episode reviews a study showing that 53BP1-RIF1 and DNA-PKcs have different genetic relationships across blunt end joining, deletion patterns, HDR, and radiosensitivity. Key terms: 53BP1, DNA-PKcs, RIF1, non-homologous end joining, microhomology.

Study Highlights:
Using EJ7-GFP and MA-del reporters in HEK293 cells, loss of 53BP1 alone did not reduce blunt No Indel EJ but amplified the decrease caused by DNA-PKcs kinase inhibition (M3814) or PRKDC knockout. Disruption of 53BP1 or RIF1, and DNA-PKcs inhibition or loss, each caused a similar shift toward deletions with increased microhomology and reduced non-microhomology deletions, with combined disruption not additive. 53BP1 loss reduced blunt EJ efficiency in XLF-deficient cells and the deletion pattern of the 53BP1/XLF double mutant resembled that of 53BP1 loss alone. DNA-PKcs kinase inhibition produced marked increases in HDR and radiosensitivity that differed from genetic DNA-PKcs loss and were not fully additive with 53BP1 or RIF1 loss.

Conclusion:
53BP1-RIF1 act as a backup to DNA-PKcs for blunt DSB end joining but function in the same pathway as DNA-PKcs to limit microhomology-mediated deletions, while DNA-PKcs kinase inhibition broadly perturbs repair outcomes and increases radiosensitivity

Music:
Enjoy the music based on this article at the end of the episode.

First author:
Makins K

Journal:
Nature Communications

DOI:
10.1038/s41467-025-65329-3

Reference:
Makins K, Cisneros-Aguirre M, Lopezcolorado FW & Stark JM. 53BP1-RIF1 and DNA-PKcs show distinct genetic interactions with diverse chromosomal break repair outcomes. Nature Communications. 2025;16:10361. https://doi.org/10.1038/s41467-025-65329-3

License:
This episode is based on an open-access article published under the Creative Commons Attribution 4.0 International License (CC BY 4.0) – https://creativecommons.org/licenses/by/4.0/

Support:
Base by Base – Stripe donations: https://donate.stripe.com/7sY4gz71B2sN3RWac5gEg00

Official website https://basebybase.com

On PaperCast Base by Base you’ll discover the latest in genomics, functional genomics, structural genomics, and proteomics.

Episode link: https://basebybase.com/episodes/53bp1-rif1-dna-pkcs

QC:
This episode was checked against the original article PDF and publication metadata for the episode release published on 2025-12-02.

QC Scope:
- article metadata and core scientific claims from the narration
- excludes analogies, intro/outro, and music
- transcript coverage: Audited the transcript sections describing (a) the interplay between 53BP1-RIF1 and DNA-PKcs in blunt end joining, (b) deletion/microhomology patterns (MA-del and related assays), (c) HDR and radiosensitivity outcomes (LMNA-HDR and clonogenic assays), (d) role of XLF and RIF1 as context modifiers, and (e) comparisons b
- transcript topics: EJ7-GFP No Indel EJ assay and 53BP1/DNA-PKcs interaction; MA-del assay: deletion patterns and microhomology usage; HDR frequency via LMNA-HDR assay; Radiosensitivity and clonogenic survival after IR; Roles of 53BP1, RIF1, XLF in end-joining and synapsis; DNA-PKcs kinase inhibition vs genetic loss

QC Summary:
- factual score: 10/10
- metadata score: 10/10
- supported core claims: 7
- claims flagged for review: 0
- metadata checks passed: 3
- metadata issues found: 0

Metadata Audited:
- article_doi
- article_title
- article_journal
- license

Factual Items Audited:
- 53BP1-RIF1 acts as a backup to DNA-PKcs for blunt DSB end joining (NHEJ).
- Disruption of 53BP1 or RIF1 increases microhomology-mediated deletions; combined disruption with DNA-PKcs disruption is not additive for this outcome.
- DNA-PKcs kinase inhibition (M3814) produces larger shifts in repair outcomes (HDR increases and radiosensitivity) than genetic loss of DNA-PKcs.
- HDR frequency increases with 53BP1 loss in DNA-PKcs-proficient cells; M3814 treatment also increases HDR; combined perturbations show context-dependent effects.
- XLF interacts with 53BP1 in promoting end-joining; 53BP1 loss can reduce blunt EJ in XLF-deficient cells.
- RIF1 loss mirrors 53BP1 loss in several microhomology deletion patterns and its effects become pronounced under DNA-PKcs inhibition.

QC result: Pass.